This invention concerns methods for denaturing double-stranded plasmid DNAs.
It is common in the art to determine the nucleotide base sequence of DNA molecules. One type of DNA molecule used in such procedures is supercoiled plasmid DNA. Such DNA must be denatured in some manner prior to use in a chain-termination sequencing reaction. Generally, the DNA is denatured by treating with 0.2M NaOH, and then ethanol precipitating the denatured strands so that they may be used in a sequencing protocol. Denaturation by simple heating is not generally possible since the melting temperature of covalently closed circular DNAs is above the boiling point (100.degree. C., see below). Common protocols are described by Tabor and Richardson, U.S. Pat. Nos. 4,795,699 and 4,962,020.